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Establishing a Top-Down Proteomics Platform on a Time-of-Flight Instrument with Electron-Activated Dissociation

ABSTRACT: . Top-down proteomics is the study of intact proteins and their post-translational modifications with mass spectrometry. Historically, this field is more challenging than its bottom-up counterpart because the species are much bigger and have a larger number of possible combinations of sequences and modifications, thus there is a great need for technological development. With improvements in instrumentation and a multiplicity of fragmentation modes available, top-down proteomics is quickly gaining in popularity with renewed attention on increasing confidence in identification and quantification. Here, we systematically evaluated the Sciex ZenoTOF 7600 system for top-down proteomics, applying standards in the field to validate the platform and further experimenting with its capabilities in electron-activated dissociation and post-translational modification site localization. The instrument demonstrated robustness in standard proteins for platform QC, as aided by zeno-trapping. We were also able to apply this to histone post-translational modifications, achieving high sequence coverage that allowed PTMs site-localization across protein sequences with optimized EAD fragmentation. We demonstrated the ability to analyze proteins spanning the mass range and included analysis of glycosylated proteins. This is a reference point for future top-down proteomics experiments to be conducted on the ZenoTOF 7600 system.

INSTRUMENT(S): ZenoTOF 7600

ORGANISM(S): Bovinae (ncbitaxon:27592)

SUBMITTER: Benjamin A. Garcia

PROVIDER: MSV000095995 | MassIVE |

SECONDARY ACCESSION(S): PXD056407

REPOSITORIES: MassIVE

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